Fig. 5: Functional analysis of three candidate genes.

Allele of CsANRa: CGT; Allele of CsANRb: GCC; Allele of CsF3′5′Ha: TGTT; Allele of CsF3′5′Hb: TCAC; Allele of CsMYB5a: CAA; Allele of CsMYB5b: CTA. a Transient overexpression of these three genes was conducted in tobacco leaves in which secondary metabolites were subsequently detected by LC–MS. The metabolite profiling of all the overexpression tobacco lines is different from that of the GFP control. The metabolite profiling of the two alleles of each gene is distinct from each other. b The purified enzymes were incubated with tea extracts revealing their catalytic function in tea secondary metabolism. The dots with the same color in (a) and (b) indicate the four biological replicates for metabolite profiling. c Heatmap of significantly changed metabolites in both tobacco leaves (left) and tea extract (right) with presence of CsANR (P < 0.05; t test; two sided). The free GFP was used as negative control. d Enzyme activity of the anthocyanidin reductase (ANR). The Arabidopsis ANR (AtANR) was used as assay control. Four measurements were conducted for each enzyme under each concentration of cyanidin chloride. Data points are jittered to improve legibility. Shadows indicate the 95% confidence interval of linear regression. Two independent experiments were performed and generated similar results. Source data underlying a, b, d are provided as a Source Data file.